The invention relates to a method of infrared-optically determining the concentration of at least one analyte in a liquid sample, wherein the infrared absorption of the analyte is measured and compared with a standard.
The invention further relates to a device for the infrared-optical transmission determination of the concentration of at least one analyte in a liquid sample, with a sample cuvette filled with the sample liquid, the sample cuvette being arranged in the radiation path between a radiation source for providing the infrared radiation and a detector for measuring the infrared absorption induced by the analyte in the sample cuvette.
Moreover, the invention relates to the use of such an arrangement for the infrared-optical transmission determination of the concentration of at least one analyte in a liquid sample.
The detection or measurement of concentration of substances in a sample is performed in many scientific and technological fields, e.g. chemistry, process technology, production technology, medical technology, environmental analytics, and food analytics, by means of absorption spectra. The infrared spectrum is particularly suitable, since precisely in this range many analytes have characteristic absorption bands from the intensity of which the analyte concentration can be determined.
GB 1 521 085 A discloses a detector for an infrared analyzer, which serves to determine the concentration of a certain component in a liquid or gaseous sample. A filter that passes infrared radiation of a single narrow wavelength range is placed between the sample cuvette and the radiation source. A wavelength range is chosen which is absorbed by the substance to be analyzed. By the difference between the absorption spectra of the sample with analyte and the sample without analyte, the presence as well as the concentration of the analyte in the sample can be determined. This described analyzer, however, requires a complicated arrangement, and the results obtained are not sufficiently specific.
WO 921/7767 relates to a method for quantitating fat in a fat-particle-containing emulsion by using infrared absorption techniques, wherein the absorption peak at a wavenumber of approximately 1160-1190 cmxe2x88x921 is used for determining the fat concentration.
AT 404,514 B describes a further arrangement for measuring analytes in a liquid sample by infrared absorption. Before the measurement, the analyte to be measured is subjected to a chemical reaction which leaves all the remaining components of the liquid sample unaffected, and the change in the infrared absorption caused by the chemical reaction with the analyte is measured as a function of the analyte concentration to be determined. This chemical reaction is, e.g., a change of the pH, so that the substance to be analyzed is present in a certain form before the pH change, such as a single-charged substrate or an uncharged phosphoric acid which is non-absorbing or only slightly absorbing at the wavelength indicated. After the pH change, the analyte is present in a form, e.g. triple-charged phosphate, which has an absorption maximum at the wavelength indicated. From the difference of measurement before and after the chemical reaction, the presence and the concentration of the analyte can be determined. To produce light with a certain wavelength, a filter that passes infrared radiation of a single narrow wavelength range is placed between the sample cuvette and the source of radiation. This method is simple and rapid, yet in view of the described chemical reaction, includes difficulties as regards the sensitivity and robustness of the analyzer.
It also is known in the art that a selective concentration determination of glucose in complex mixtures such as human serum, is possible by absorption measurement at a few wavelengths in the middle region of the infrared spectrum, as described by Heise et al. in Fresenius J. Anal. Chem. (1997), 359, 93-99. Absorption spectra in the middle of the infrared spectrum were taken on blood plasma and whole blood samples using a Fourier Transform Infrared (FT-IR) spectrometer. Heise et al. also disclosed a chemometric model for the determination of glucose in unknown samples, where a few wavelengths were sufficient to obtain results equal to, or better than, results obtained by means of a PLS (partial least square) model covering the entire (1200-950 cmxe2x88x921) spectral range. A drawback of this method is, however, that the FT-IR spectrometer is not handy and is heavy and the employed measurement on the surface of a toxic ZnSe crystal is not usable for on-line determinations of biological samples in so far as the samples, due to the contact with the toxic ZnSe crystal, also become toxic after the determination. Moreover, there are potential problems with the absorption of proteins on the surface, and, lastly, transmission measurements in practice cannot be made since the FT-IR spectrometer, which has a low light intensity, only allows for the use of layers only up to 50 micrometers thick. Such layer thicknesses, however, are not suitable particularly when determining the concentration in biological samples, since within a short time the thin layer will be clogged or the sample (e.g. whole blood) will be damaged, whereby an on-line measurement, e.g. on the living patient, with a return of the sample to the patient becomes life-threatening and thus completely impossible.
In general, conventional radiation sources for producing infrared radiation are based on thermic radiators and accordingly are limited in their radiation power. Thus, the radiation power emitted by a thermic radiator of 1500 K in the narrow spectral range of from 9.9 to 10.1 cmxe2x88x921 is less than 0.2% of the entire radiation power emitted. Due to the practical difficulties of efficiently collecting emitted radiation, only a small fraction of the emitted radiation is actually available for the measurement. For example, a state of the art spectrometer from Bruker GmbH, the radiation power distributed over the entire region of the spectrum and finally available in the sample chamber is only approximately 25 mW. The consequence thereof is that in the narrow wavelength range from 9.9 to 10.1 cmxe2x88x921, only low power, approximately 50 xcexcW, is available. With respect to the thermic radiator as disclosed in AT 404,514, it is estimated that the usable power is only in the range of approximately 100 xcexcW.
Thus, it is an object of the present invention to provide a method as well as an arrangement for the infrared-optic transmission determination of the concentration of an analyte in a liquid sample, which can be carried out easily and rapidly, has a sufficient sensitivity and robustness so as to afford a marketable product, and which, in particular, is suitable for on-line measurement of biological samples where the sample is returned to the patient.
The method is characterized in that the liquid sample is irradiated with infrared radiation of high light density, wherein the infrared radiation has a sharp intensity peak at at least one wavelength.
Such infrared radiation with a high power density is produced with a laser, wherein, due to the high spectral density and the sharp intensity peaks with lasers, spectroscopic methods are possible, which cannot be carried out with conventional radiation sources. The combination of the [infrared] spectroscopy with laser spectroscopy results in a method for infrared-optic transmission determination of analytes in a liquid sample, which has a markedly higher sensitivity, flexibility and robustness than conventional methods.
With the method according to the invention, concentrations of ions as well as any other substances, such as organic acids, in particular fatty acids, alcohols, carbohydrates, in particular glucose, proteins, urea etc. can be measured with high precision.